Neurodegenerative disorders with prominent filamentous alpha-synuclein (alpha-syn) aggregates in CNS neurons or glia are known as synucleinopathies. Synucleinopathies characterized by neuronal alpha-syn inclusions, or Lewy bodies, include Parkinson's disease (PD) and dementia with LBs (DLB), while multiple system atrophy (MSA) is characterized by numerous glial cell inclusions (GCIs) and less frequent LB-like inclusions formed by fibrillar alpha-syn. Most PD is sporadic, but mutations/duplications in the alpha-syn gene of rare kindreds cause autosomal dominant hereditary PD/DLB. Thus, filamentous alpha-syn inclusions are linked to the onset/progression of synucleinopathies. In the current funding cycle, transgenic (TG) mouse models of synucleinopathies were generated that developed neuronal and glial alpha-syn pathologies recapilulating those in DLB and MSA, respectively. We now propose to use these existing TG mice together with new TG mice we will generate to test specific hypotheses regarding toxicity of a-syn and the mechanisms of alpha-syn aggregate formation. In Aim 1, we will cross our MSA-model TG mice with glial pathology, with those TG mice expressing alpha-syn in neurons, to test the hypothesis that oligodendrocyte degeneration enhances neurodegeneration. In Aim 2, we will address the significance of dopamine interactions with alpha-syn. Since all TG mice reported to express alpha-syn in neurons do not develop inclusions in dopaminergic substantia nigra (SN) neurons, we hypothesize this is due to dopamine mediated inhibition of alpha-syn fibrillization through interactions with residues 125-127. Now, we propose to test this hypothesis by generating TG mice expressing alpha-syn with these residues mutagenized asking if the removal of the dopamine/alpha-syn interaction facilitates formation of SN alpha-syn neuronal inclusions. In Aims 3, 4 and 5, we will test the hypotheses that the C-terminus of alpha-syn and Ser129 phosphorylation play important roles in the pathogenesis of alpha-syn lesions and that these lesions induce neurodegeneration in TG by impairing axonal transport. Taken together, our work will lead to more informative models of synucleinopathies and address important questions on mechanisms leading to alpha-syn pathologies and their contribution to brain degeneration in synucleinopathies.